1. Field of the Invention
Some diseases are caused by an insufficiency in the number of certain types of cells which died for various reasons. In cirrhosis of the liver, a lack of hepatocytes in sufficient number to detoxify accumulating metaholic poisons causes coma. In Addison's disease, a lack of adrenal cortical cells causes death unless the diseased individual receives steroid hormones that are usually provided by these cortical cells. In Type I diabetes mellitus, a lack of pancreatic islet beta cells causes death unless the diseased individual receives insulin that is usually provided by these beta cells.
Sometimes, an insufficiency in the number of certain cells is not caused by disease but is the result of an adverse side effect to chemotherapy of diseases like cancer. Drugs and radiation used to kill cancer cells also kill normal hemopoietic marrow and lymphopoietic cells and patients die from the lack of blood cells.
Unfortunately, unlike the lower forms such as the planarian worms, mammals can not readily grow replacement cells because, generally, mature, differentiated cells do not self-reproduce much if at all. Mammals also reject any transplant of replacement cells if such cells are taken from anyone else than self. (One exception to this rule is where cells are transplanted between identical twins (syngeneic transplant)). Rejection occurs because, like fingerprints, every individual has a unique histocompatibility protein called transplantation antigen on the membrane surface of his/her cells. Every individual also has immunologic defense cells called lymphocytes which are capable of recognizing as foreign any cells from other individuals. Any transplanted cells possessing foreign transplantation antigens are promptly rejected and killed by these lymphocytes. The uniqueness of each individual's antigen is the result of the uniqueness of the individual's desoxyribonucleic acid (DNA). The transplantation antigen is made employing a messenger ribonucleic acid (mRNA) template which in turn is a complementary copy of the DNA's histocompatibility locus. With the exceptions of identical twins and members of a closely inbred strains of animals, who are genetically identical (i.e., possess identical DNA's), everyone else has his/her own unique code at the histocompatibility locus.
Since at this time it is not possible to make mature, differentiated cells to replicate themselves in order to replenish their number, and since transplanting replacement cells from other individuals is fraught with much difficulty and risks, I thought that it could be desirable to construct a new cell which: 1) possessed transplantation antigenicity compatible with the diseased or deficient individual, and 2) possessed function of the cells whose number is lacking in the diseased or deficient individual. A large number of such cells can then be successfully transplanted into the diseased or deficient individual and correct the insufficiency.
In setting about on the task of constructing a new cell with above requirements, I relied on the following hypotheses.
A. The specificity of the transplantation antigen on the cell surfaces is determined by the nuclear DNA.
B. Except for the germ cells, nuclear DNA's of every cell in the body, whether from skin epithelium, mesenchymal fibroblast, intestine, myocyte, hepatocyte, or pancreatic islet, all contain the same genetic code.
C. Although the DNA's of different tissues are identical, different parts of the DNA are being expressed and so different proteins are being caused to be synthesized; different proteins make one cell to be, for example, myotube and another to be hepatocyte.
D. There can be present in the cytoplasm, factors which direct adjacent nuclear DNA to continue the expression that differentiates the cell as, for example, kidney or liver; similarly, there can be present in the cytoplasm, factors that encourage DNA to frequently enter the mitotic cycle, i.e., making the cell self-replicating.
2. Description of the Prior Art
Constructing a new cell with new properties has become possible owing to the technology of fusing two whole cells or fusing nucleus (karyoplast) of one cell with cytoplasm (cytoplast) of another cell. I believe the credit for pioneering in this art goes to Henry Harris (1965. Nature, London, 206: 583-1966. J. Cell Sci. 1: 1-1966. Proc. Roy. Soc. Brit. 166: 358-) and to D. M. Prescott (1974. Proc. Natl. Acad. Sci. 71(5): 1999-2002). An improved method for fusing cells and their components using polyethylene glycol (PEG) has been described by R. L. Davidson (1978. Natl. Cancer Inst. Monofr. 48: 21-30). Some of the hypotheses described in the preceding paragraph on which this invention is predicated have been tested and investigated by various workers (R. L. Davidson, cited above; Choy-Pik Chiu and Helen Blau, 1984. Cell 37: 879-887; Inge Schwag and O. Luger, 1980. Differentiation 16: 93-99). Their work gave me greater confidence that my ideas would work. However, I had no assurance of success without actually practicing my methods because there are contradicting findings among the work just cited and in the rest of the literature on the mechanisms of differentiation and cell proliferation.